Lung cancer remains the most common cause of death from cancer in the US and there is a high need for improved therapeutic interventions. Non-Small Cell Lung Cancer (NSCLC) which accounts for 85% of all lung cancer cases, has a poor 5 year survival rate of only 16.8%. Epidermal growth factor receptor (EGFR) is a major target for the treatment of (NSCLC), but only a subset of patients, those who have EGFR mutations, benefit from EGFR tyrosine kinase inhibitors (TKIs) such as erlotinib and afatinib (FDA approved drugs). Intrinsic lack of response to erlotinib in wild-type (WT) EGFR and development of acquired resistance in mutant EGFR NSCLC could be driven by a transition from an epithelial to a mesenchymal state, induction of a stem-cell phenotype, and other STAT3-dependent mechanisms. Critical to the development of intrinsic and acquired resistance to EGFR TKIs, is the transcription factor signal transducer and activator of transcription 3 (STAT3). STAT3 is persistently activated in 22% - 65% of NSCLC and overexpression of STAT3 is associated with poor prognosis, making it an attractive target to overcome EGFR TKIs for NSCLC therapy. STAT3 inhibitors that target STAT3 activation (by inhibiting phosphorylation) are clinically limited because they lack the ability to effectively block STAT3 dimerization and nuclear translocation. To address this problem in AIM 1, we are currently testing the ability of a novel STAT3 inhibitor (cyclic STAT3 decoy ? CS3D) and have generated preliminary evidence that CS3D produces anti-tumor effects in NSCLC independent of EGFR mutation status. CS3D acts as a STAT3 ?decoy? by mimicking the DNA consensus sequence in the promoter region of STAT3- responsive genes, causing the binding of STAT3 dimers and preventing their nuclear translocation and the transcriptional regulation of pro-survival genes. Molecular and genetic in vitro approaches will be used to demonstrate CS3D specificity for STAT3 compared to an inactive mutant control (CS3M). To further validate the anti-tumor effects of CS3D in AIM 1, we will utilize a variety of in vivo models (xenograft, orthotopic, and immune-competent) to confirm the therapeutic effects of CS3D as compared to CS3M. Assessment of biomarkers of CS3D sensitivity in WT and mutant EGFR NSCLC will provide predictive markers to identify patients that would benefit from this therapeutic approach. In AIM 2, we also predict that CS3D combined with afatinib, (an EGFR/HER2 irreversible TKI) will produce a superior anti-tumor effect compared to single agents. We will further test the ability of CS3D to re-sensitize afatinib-resistant cells to EGFR inhibition by reversing the EMT and stem-cell phenotype. Biomarkers of drug sensitivity will also be assessed in AIM 2. The rationale for this proposal is both to utilize a more effective approach to target STAT3 and to demonstrate the general principle that combination treatment has greater therapeutic benefit than single therapy for NSCLC.